Benzyl-penicilloate of tetracycline



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BENZYL-PENICELOATE F TETRACYCLINE Lee C. Cheney, Fayetteville, and William J. Gottstein, Syracuse, N. Y., assignors to Bristol Laboratories Inc., Syracuse, N. Y., a corporation of New York No Drawing. Application February 1, 1956 Serial No. 562,630

4 Claims. (Cl. 260-3065) The present invention relates to new, non-toxic waterinsoluble amine salts, more particularly to penicilloic acid salts of tetracycline, which are capable of exerting a repository antibiotic action and are also useful for oral, therapeutic administration, for external application, for use as a growth-stimulating supplement, as in animal and poultry feeds, drinking water and by implantation, and for use to stimulate the growth of plants, such as radishes, oats and grass, and for use in manufacturing procedures for isolation and purification to enable substantially quantitative removal of tetracycline from solution. This application is a continuation-in-part of our prior, (so-pending application S. N. 396,748, filed December 7, 1953, now abandoned.

The tetracycline D-benzyl-penicilloate of the present invention may be obtained by reaction of penicillin acid with tetracycline in a water-immiscible organic solvent or by the metathetical reaction in water of a Watersoluble penicillin salt and a water-soluble salt of tetracycline.

Tetracycline in the form of its free base or an acid addition salt such as the hydrochloride is prepared according to the Journal of the American Chemical Society, volume 75, pages 46214623, 1953. D-benzyypenicilloic acid is described on page 574 of The Chemistry of Penicillin: Clark, Johnson and Robinson, Princeton Press, 1949. 1 A more comprehensive understanding of this invention is obtained by reference to the following examples which are illustrative only and are not the exclusive embodiment of the invention.

Example 1 Acid penicillin G isopropyletherat'e (0.492 g.) dissolved inl0 ml. methyl isobutyl ketone was added to a solution of 0.5 g. tetracycline dissolved in ml. methyl isobutyl ketone. Solid tetracycline D-benzylpenicilloate precipitated and was collected by filtration and found to melt over the range l36-l56 C. with decomposition. Found by analysis: C, 57.6 percent; H, 5.78 percent.

The compound was active in the standard penicillin bioassay.

Example 11 Potassium penicillin G (4.4 g.) was suspended in 50 m1. methyl isobutyl ketone, acidified with dilute phosphoric acid and the organic layer was washed with- 50 United States Patent-O ml. water, dried over anhydrous sodium sulfate and added to a solution of 5 grams tetracycline in 150 ml. methyl isobutyl ketone. The solution became turbid and was concentrated on a warm water bath (40 C.) in vacuo to one-third its original volume and then cooled. Solid tetracycline D-benzylpenicilloate precipitated and was collected by filtration, slurried with ml. ether containing 1 ml. water, collected by filtration and dried in air. The product was crystalline and soluble in water to the extent of only about 0.16 percent by weight.

Example 111 Potassium benzylpenicillin (18.6 g., 0.05 mole) in 200 ml. water is mixed under cooling with a saturated solution of tetracycline hydrochloride (25.5 g., 0.05 mole) in Water. Tetracycline D-benzylpenicilloate forms and, after the volume has been reduced by distillation in vacuo, is collected by decantation or filtration and dried in vacuo.

Example IV Acid penicillin isopropyletherate (0.492 g.; a commercial mixture of the several penicillins) dissolved in 10 ml. methyl isobutyl ketone is added to a solution of 0.5 g. tetracycline dissolved in 15 m1. methyl isobutyl ketone. Solid tetracycline D-benzylpenicilloate percipitates and is collected by filtration.

Example V Two and four-tenths kg. (5 moles) of tetracycline hydrochloride were dissolved in 21 liters of water. Twohundred milliliters of concentrated hydrochloric acid were added to adjust the pH to 1.5. This solution was added to 1.965 kg. (5.23 moles) of potassium penicillin G dissolved in 10 liters of water at 0 C. Upon being stirred vigorously for /2 hour, the solution was filtered and the collected salt was washed with 1 liter of cold water and air dried to constant weight; yield 3.432 kg. (88%). M. P. 164165 C. with decomposition (softens [oz] =54.O5 (C=l, methyl Cellosolve). Tetracycline potency 728, 770, 700 meg/mg. by bioassay. No penicillin activity was found either by bioassay or chemical assay. Moisture 2.9%. Anal. calcd for C H N O C H N O S (tetracycline D-benzylpenicilloate) S, 4.02; found: S, 3.77. 1

Example VI Twenty grams of tetracycline D-benzylpenicilloate was suspended in 150 ml. water at pH 1 in an ice-bath. The mixture was extracted in the cold with five 100 ml. portions of methyl isobutyl ketone. After separation, tetracycline was recovered from the aqueous phase by presample of the disodium salt was dissolved in 5 ml; of; water. The' solutionwas'cooled'in an ice bath with a few particles of ice added directly to the salt solution. Upon acidification with 2 drops of concentrated hydrochloric acid and swirling, a white amorphous solid formed which was filtered E immediately and air dried. M. P. Ill-114 C.; [a] =+55.1 (6:.2 methanol).

Regeneration of tetracycline D-benzylpenicill0ate.- One gram of the disodium D-benzylpenicilloate was dissolved in ml. of water. To this solution was added 1.22 grams of tetracycline hydrochloride in ml, of

' .water. Two drops of concentrated hydrochloric acid were added to lower the pH to about 2. 'The solution was stirred for 15 minutes, filtered, and the salt was dried to constant weight (0.7 gm); P. 160164 decomp.; =59.12-(C,=;5 rriethyl' Cellosolve). Y

-The solubilitylofttetracycline Drbenzylpenicilloat'e in isaturatedasolution in :waterat 25 -C. was measured by ;bioassay with thetollowing results: V p

While the present invention hasvbeen described with particula'r re'ference .to -penicillinG it will be understood that other penicillins arealso included within the scope of this'invention; For instance, -penicillins such as the :penicillins G, F, X, 'dihydro F, 0,-BT, andK and mixtures of twopor more such penicillins, particularly such -niixtures containing at least 85% of penicillin G, are included within the scope of-this invention.

It will be understood that the reaction canbecarried ont" in water-immiscible organic solvents other than'ether.

tragacanth, gelatin, pectin, alginates, dextran's, guin karaya, lecithin, ,Sp'ans, Tweens, Amerchols, inositolplios phoric acids and their non-toxic salts (e, g. sodium phytate), glyceryl monostearate, Kreelon CD, polyoxyallqe' lene-sorbitols, no more than 2.0 percent of an injectable oil such as peanut oil and condensation products, having molecular weights greater than 1500, of ethylene oxide 7 with a condensation product. of propylene oxidewith propylene glycol (e. g.-Pluronic F68). On occasion, it is advantageousto actually coat the particles of salt, at least:

in part, with one of these agents, e. g. lecithin,

If desired, buffering agents are added such as sodium' citrate or sodium phosphates, as when the salts of this'in vention are used with an alkali metal salt of penicillin. When appropriate, the suspensions may be buttered by the addition of free acids, 'e; g. citric acid, phosphoric acid. If desired, a stabilizing, agent 'is added, such as sodium hexametaphosphate, hexamethylenetetramine or 7 sodium phytate. Smallamounts of, preservative are often used, e. g. phenoL cresol or alkyl esters of'p-hydrox'ybenzoic acid.. Useful variations in the properties'of the salts of the present inventio'n 'are obtained by altering particle size or shape, as' by'varying'procedu res ofcrystallization or, mosteasily, by -mechanical means such as grinding, hammer-milling;pulverizing, or micronizing,

When desired for specific purposes, administered by appropriate routes, and rendered pharmaceutically' compatible as described in the art for water-insoluble salts of penicillin, there may be admixed with thefsalts :of the present invention, and particularlydhose, preparedfor;

oral use, one or more ofvarious othermedicaments such it ,as antihistamines,-sulfa drugs (e. .g. sulfadiazine, sulfa- Examples of such solvents are butanol, amyl acetate,

methyl jamyl acetate, isopropylether, mesityl oxide,

methyl isobutyl ketone, methylene dichloride,- ethylene dichloride and chloroform; Recovery, of' the product 7 maybe increased by' evap rating the solvent in vacuo at low temperatures V It willbe understood also thatthe solution in an organic solventjof'the ifree amine mayfbe prepared in the organic solvent ,by theuse of caustic .to liberate the free amine from janbrg'anic-solvent soluble or water soluble salt su'chj asj-theiliydr'ochloride, phosphate, nitrate, hydrofbfomide flsulfate, citrate, acetate and tartrate.

merazine, sulfamethazine, sulfac'etarnide, sulfanilamide,

' sulfapyridine, sulfathiazole, sulfapyrazine, 'sulfaguanidine, sulfathalidine, sulfit'suxidine, 'sulfisoxazole, sulfarnylon,

phthalylsulfacetamide, N3,4-dimethylbenzoyhsulfanilamide, benzylsulfanilamide and N'-(2-quinoxalyl)-sulfanilamide), vitamins (e. g. vitamins A, B li B B and members of thatfamily, folicacid and members of that family, vitamins C,'D D and E),"lipotro'pic agents, stimulants (e. g.'caifeine,amphetamines), analgesics(e; g'. aspirin, s'alicylamide, sodium gentisate, p-acetylaminoe phenol, phenacetin, codeine), laxatives (e. g. phenolphthalein),-sedatives' (e. g. barbiturates, bromides), salts of penicillin (e. g.- potassium penicillin G, procaine peni- .cillin G, 1-ephenamine penicillin G,'dibenzylamine peni- Thefsalts of the presentinven tion, whether aloneoriin 'conib'inationwith other therapeutic agents, are used asfa simplepowder, as tablets, as-troches, as lozenges, or in capsules. ,Aqueousf'suspensions are preparedin advance or'insitu when the stability of other added agents'(e. g.

pota'ss'iumbenzylpenicillin) so requires, andare a highly V [acceptable oral .dosageformx A particularly useful oral dosage' form is a'suspension ina2-palatable, acceptable edible oil; coconut oil is preferred, particularly when fitee of*tristear'inand gelled withan1aluminum stearate (cf UQS. Patent 2,507,193). The salts of the present invention are admixed with'suitable .b'uflers or pharmaceutical additives, ,preservatives, diluents, binders, lubricants, mas- The salts 'offlthe. present invention, alone or in cornbi nation with other therapeutic agents, arepresented for tparenteraljuse as a'suspension in an injectahle oilte. .g.

peanut oil), I as 'a suspension in a gelled injectableoil (e. g. peanut oil gelled with.2% aluminum monostearate), as an aqueous suspension oras'a powder to which a sterile, aqueous diluent is addedlhefore use togenerate an injec'table'suspension. These aqueoussuspensions often V advantageously 'containnon-toxic suspending. or dispersing agents, suchas sodium ,carboxymethylcellulose, methylcellulose, polyvinyl alcohol, 'polyvinyl-pyrrolidone," gum 'ticatorysubstances,colors, flavors, suspending'agents, dis- *persingagents,resuspending agents, and stabilizing agents biotic agents (e. 'g. -neomycin, bacitracin, polymixin,tyrothricin, erythromy V cin, Aure'omyc in, Terramycimtetracycline; chloramplienr' icol,magnamycin; in some .cases such combinations gattack awider range ofio'rganismslor show synergistic efllcacy or providedec'reased toxicity with equal etncacy) cillin G, other salts "disclosed by U. S. Patent 2,627,491;

these combinations are particularly useful to enable var-.

iation of the pattern of blood levels obtained) other antis'trcptomycin, dihydrostreptornycin,

and spreading agents (e.-g. hyaluronidase). Multiple combinations ofsuch'adde'd'agents are often useful, e. g. sulfadia'zine and sulfamerazine to provide". equal efiicacy and reduced toxicity; substantially equal, amounts of a non-toxic acid. additionsalt'of-istrepto-' mycin and anon-toxic acid addition salt of .dihydrostreptomycin (sulfate; sulfate iodine) to provide equal Jefficacy and reduced toxicity; jaspirin, .phenacejtin and caffeine; multiple vitamins;i h'ighly'water-soluble salts of penicillin (e.' g'. potassium) withmoderately water-insoluble salts of penicillin (e..g. procaine).

Further information which is; readily applied to formulations and 'combinations of the' tsalts of the present .invention is to befo'und in U S. Patents 2,602,038, 2,608,507,

2,608,509, 2,619,447, and 2,578,651.

-We claim: 7 1. Tetracycline. penicilloate. 2. Tetracycline Dsben'zylpenicilloate. a 3; The process of mixing in the coldan aqueous solution of tetracycline below pH 2 with a substantially equivalent quantity of a water-soluble salt of penicillin G to precipitate tetracycline D-benzylpenicilloate.

4. The process of mixing in the cold an aqueous solution of tetracycline below pH 2 with a substantially equivalent quantity of potassium penicillin G to precipitate tetracycline D-benzylpenicilloate.

References Cited in the file of this patent UNITED STATES PATENTS 2,627,491 Szabo et a1. Feb. 3, 1953 FOREIGN PATENTS Belgium Sept. 29, 1951 OTHER REFERENCES Drug Trade News, vol. 26, Jan. 22, 1951 p. 49 (Mfg. section).

American J. of Pharmacy, July 1945, p. 253.

Price et 211.: Annals of the N. Y. Acad. Sci, vol. 51, Art. 2, Nov. 30, 1948, p. 216. 

1. TETRACYCLINE PENICILLOATE. 